ES2471877T3 - A composition for use in the treatment of lactose intolerance or conditions resulting from lactase deficiency - Google Patents

Abstract

A composition for use in the therapy of lactose intolerance or conditions resulting from lactase deficiency, wherein the composition is a solid non-dairy dosage form comprising: (a) a lactase which is stable and active in a pH range of 3 to 6; the term stable means that lactase (a) still has at least 30% of its activity after one hour at 37 ° C at a pH in the range of 3 to 6; and (b) one or more microorganisms that produce lactase and / or that contain lactase selected from Lactobacillus delbrueckii ssp bulgaricus and Steptococcus thermophilus; wherein the lactase (a) is different from the lactase derived from said one or more microorganisms (b); and wherein the one or more microorganisms that produce lactase and / or that contain lactase (b) are in dry form.

Description

A composition for use in the treatment of lactose intolerance or conditions resulting from lactase deficiency

The subject matter of the invention is an agent for use in administration to subjects suffering from lactose intolerance, which contains the enzyme lactase (without. Tilactase, tilactasum, beta-galactosidase) in combination with microorganisms. In the context of this patent application, the term lactose intolerance is used to refer not only to lactose intolerance and medically defined lactose metabolism disorder (lactose malabsorption, poor digestion of the lactose), but to any form of health damage and symptoms that occur as a result of the intake of lactose or food products containing lactose, or due to the release of lactose in the digestive tract of humans or animals of other substances, such as pharmaceutical compositions.

Background of the invention

Lactose is a disaccharide and is an important nutritional component that supplies energy. It is presented as a component mainly in milk and dairy products, but it is also used by the food industry as a sweetener, filler and consistency improver, for example, in sweets, processed meats, bakery products and prepared foods. Pharmaceutical compositions often also contain lactose. In the body, lactose is cleaved into the usable nutrients glucose and galactose in the small intestine by the endogenous lactase enzyme. For babies, the lactose contained in breast milk is a very important food.

Lactose intolerance is usually the result of lactase deficiency. If insufficient amounts of the enzyme lactase are available in the body (lactase deficiency), lactose passes in an uncleaved way to the lower parts of the intestines (large intestine), where it is fermented by intestinal bacteria, causing, for example , that gases and / or an increase in the entry of water into the large intestine occur. This can lead to discomfort such as stomach ache, flatulence, bloating or diarrhea, after consumption of dairy products. This is called lactose intolerance (intolerance to milk sugar). Due to the similarity of discomfort, lactose intolerance is often confused with irritable bowel syndrome (irritable bowel, spastic colon).

Three different forms of lactase deficiency are generally recognized:

one.

 Primary Lactase Deficiency

2.

 Secondary Lactase Deficiency

3.

 Congenital lactase deficiency

The amount of lactase in the small intestine is higher in babies during the period of breastfeeding and then decreases continuously in most people for genetic reasons. The so-called resulting primary lactase deficiency is therefore the result of a normal aging process and can be observed in the vast majority (70% - 90%) of the world's adult population. Thus, almost all population groups in Africa and Asia do not tolerate lactose. But also in Germany, about 15% of adults have a primary lactase deficiency. Lactose intolerance resulting from primary lactase deficiency is then often referred to as primary lactose intolerance.

Various intestinal diseases can lead to the so-called secondary lactase deficiency (without acquired lactase deficiency), for example, Crohn's disease, ulcerative colitis, celiac disease (endemic sprouts) and other intestinal inflammations (for example, due to to viral or bacterial infections). In addition, after operations in the intestinal tract, especially after removing parts of the small intestine, the appearance of secondary lactase deficiency is not uncommon. This is called secondary or transient lactose intolerance. Secondary lactase deficiency is usually reversible after the intestinal disease that caused it has been cured.

In congenital lactase deficiency that occurs very rarely, the newborn lacks the gene responsible for the production of lactase. This leads to the total inability of the body to produce the enzyme. In such babies, strict nutrition without lactose should be maintained.

Within the meaning of this patent application, the term lactase deficiency is used to refer not only to the medically defined lactase deficiency, but to any type of lactase deficiency.

Not all changes in lactose metabolism necessarily lead to severe lactose intolerance. However, discomfort can also be observed in mild disorders of lactose metabolism. Until now, it has only been possible to avoid the discomforts mentioned above by maintaining a low lactose diet or

lactose free or taking preparations containing the enzyme lactase when eating foods containing lactose. It is not satisfactory to maintain a low lactose or lactose-free diet and, due to the valuable nutrients and vital substances contained in dairy products, it is also unfavorable from a physiological nutritional point of view. The choice of lactose-free dairy products is limited and, apart from this, they have a sweeter taste than lactose-containing dairy products, because during production lactose has already been cleaved by the addition of lactase. Many consumers do not like the taste of these products. Additionally, there are many foods in which one would not expect them to contain lactose. This means that the purchase of lactose-free foods often causes problems. Timely use of lactase preparations represents a good alternative and often significantly improves the quality of life of those affected. The amount of lactase required individually depends, on the one hand, on the user's sensitivity to lactose. This is mainly determined by the residual activity of the endogenous lactase. Apart from this, the amount of lactase required depends on the type of food, the total amount of food consumed and the amount of lactose it contains. It is known that lactase preparations are not always effective or equally effective in all affected people.

A series of products have been proposed for use in the treatment or relief of symptoms of lactose intolerance or lactase deficiency.

RU 292125 describes combinations of ground apple extract (pataca), a lactase and a fermented milk base containing S. thermophilus and L. bulgaricus. The product is intended for consumption by people suffering from lactose intolerance or lactase deficiency and the purpose of lactase seems to be to reduce the lactose content of the milk base. There is no suggestion that the two microorganisms have any beneficial effect on lactose intolerance. In addition, it is considered that the two microorganisms must be present in the 1: 4 ratio of S. thermophilus: L. bulgaricus in order to provide the best flavor and consistency.

DE2725731 (equivalent to US4034115) describes a four-stage process for the preparation of a fermented milk product that is low in lactose and is therefore suitable for consumption by people suffering from lactose intolerance. It is not intended to provide a general means of preventing or treating lactose intolerance or the symptoms of lactase deficiency that may be derived from lactose in other types of foods. The process comprises a first stage of fermentation using lactic acid bacteria (not S. thermophilus or

L. bulgaricus) to give a butter of milk; a second stage of fermentation using yogurt bacteria such as S. thermophilus or L. bulgaricus; a third stage that involves the addition of a small amount of colostrum; and a fourth stage that involves fermentation with L. acidophilus or L. bifidus.

WO2009 / 056979 (Danisco) describes among other dairy products (yogurts) that contain S. thermophilus, L. bulgaricus and exogenous lactase. The products are not intended to treat or prevent lactose intolerance or lactase deficiency although page 11 line 4 refers to the known use of lactase in the treatment of lactose intolerance.

WO2004 / 089290 (Pharmachem) describes the administration of chewable confectionery products containing a combination of probiotic microorganisms and lactase for a lactose intolerant person. The list of preferred microorganisms includes S. thermophilus but not L. bulgaricus. The purpose of lactase (see page 16) seems to be to eliminate any lactose present in confectionery products. WO2004 / 089290 does not contain examples of confectionery products containing lactase and / or a microorganism such as S. thermophilus or L. bulgaricus and there is no description as to how the microorganisms are presented. On pages 2 and 3 of WO2004 / 089290, it is suggested that the use of lyophilized microorganisms is problematic and in other places in many places it is considered that confectionery products are prepared without heating and dehydration, which suggests that way microorganisms are not used in a dry form.

US2008 / 0187525 (Porubcan) describes combinations of probiotic microorganisms and lactogenic enzymes. Lactogenic enzymes (including lactase) are intended to promote the growth of probiotic microorganisms. The combinations (or their components) can be presented in capsule form. The microorganisms may be in lyophilized form. Specific combinations of lactase with either S. thermophilus or L. bulgaricus are also not described or exemplified. This document is not related to the prophylaxis or treatment of lactose intolerance or lactase deficiency.

DE69505374 (= WO95 / 34292) describes microgranules containing various bioactive substances such as lactase and microorganisms, where the microgranules are enteric coated. The coated microgranules are intended to be incorporated into foods such as dairy products. S. thermophilus and L. bulgaricus are mentioned as examples of microorganisms. Lactose intolerance is discussed on page 3 and lactase is mentioned. There are examples of coated granules containing lactase and there are also examples of coated granules containing L. acidophilus. However, there are no descriptions of the use of lactase and S. thermophilus or L. bulgaricus as a whole and there are no examples of coated granules containing S. thermophilus or L. bulgaricus.

EP1112002 describes milk powders for administration to animals. Milk powders contain lactase that is added to prevent symptoms of lactose intolerance in animals. The powders may also contain probiotic microorganisms (see page 3). S. thermophilus is among the list of microorganisms, but L.

bulgaricus is not there. However, there are no examples of milk powders containing probiotic microorganisms as well as lactase.

WO96 / 06924 (= DE 69535290) describes a process for producing a fermented product rich in galactooligosacides and �-galactosidase (lactase) by fermentation of a medium containing S. thermophilus. On page 12, the products are considered to be tolerated by people suffering from lactase deficiency. There does not seem to be any description of the addition of exogenous lactase and it does not seem to suggest that the product can be used to prevent

or treat lactose intolerance or lactase deficiency.

US2008 / 0213320 (Eisenstein Mayer) describes compositions (which may be among other tablets or capsules) for the treatment of gastroesophageal reflux disorder that contains at least two of (1) a digestive enzyme, (2) a probiotic, and (3) stevia. Lactase is on the list of digestive enzymes. S. thermophilus and L. bulgaricus are listed among probiotics but are not considered as preferred. An example is provided and this contains lactase as well as 7 other digestive enzymes but also contains 6 probiotic species, which do not include S. thermophilus and L. bulgaricus. There is no reference to the treatment or prevention of lactose intolerance or lactase deficiency.

US2009 / 110674 (Loizou Nicos) describes natural food supplements that contain more than fifty different active components selected from among other minerals, enzymes, vitamins, plant extracts, selected probiotics and amino acids. On page 3, a composition is described that contains more than fifty ingredients that include lactase (5 mg) and Lactobacillus bulgaricus (2.5mg that includes Bifidus). On page 12, lactase is described as essential for the hydrolysis of lactose in milk and an enzyme deficiency is considered to cause lactose intolerance. The compositions are described as a universal panacea that encompasses general health and are not directed at any state or condition of a disease.

US2008 / 0260708 refers to compositions to "normalize the chemistry of the human body" and is intended as a universal panacea to promote good health. The compositions must contain among others a digestive enzyme, soluble and insoluble fiber, a laxative, one or more probiotics, protease, lipase, various minerals and various extracts of grass and plants. The compositions are not specifically aimed at preventing or treating lactose intolerance or the symptoms of lactase deficiency. Digestive enzymes may include lactase and probiotic enzymes may include Lactobacillus bulgaricus and Streptococcus thermophilus.

US6008027 (Langner) describes capsules containing microorganisms that produce lactase for the treatment of lactase deficiency. Although L. bulgaricus and S. thermophilus are mentioned, the exemplified compositions contain only L. acidophilus. There is no description in US6008027 of the inclusion of exogenous lactase. An essential feature of Langner capsules is that they are coated in an enteric polymer that prevents release in acidic stomach conditions but that breaks down to release microorganisms in the intestines.

US6410018, US6428786, US6562338 and US6562339 (all of Eisenhardt et al.) Each describe compositions for the treatment of lactose intolerance comprising a first lactase enzyme having an optimum working pH in the range of 3 to 6 and a second enzyme Lactase having an optimum pH in the range of 6 to 8. The second lactase enzyme has an enteric coating in order to prevent premature degradation in the stomach. None of the documents suggest the use of lactic acid bacteria in the compositions, methods and processes described therein.

Guarner et al. Brit. J. Nutrition, 93 (2005), 783-786 reveal that S. thermophilus and L. bulgaricus yogurt bacteria have a high lactase activity and that yogurt improves lactose digestion and eliminates symptoms of lactose intolerance. However, there is no suggestion in Guarner and others that exogenous lactase can be added

or that doing so would result in some benefit.

In addition, there are no evidences in Guarner and others that indicate that consuming yogurts that contain S. thermophilus and L. bulgaricus would have some effect on the digestion of lactose in subjects that poorly absorb lactose that other dairy products have consumed such as Unfermented milk and ice cream.

Although eating yogurt can provide general benefits to some who poorly absorb lactose who enjoy eating yogurt, many people do not like the taste and texture of yogurt. Therefore, yogurt containing live cultures of bacteria such as S. thermophilus and L. bulgaricus does not provide a general solution to the problem of lactose malabsorption and lactose intolerance.

Guarner and others (see above) emphasize the importance of the use of live cultures in order to produce beneficial effects. If yogurt is treated with heat, the benefits are lost. Guarner and others do not provide evidence to suggest that the administration of inactive bacteria (for example lyophilized bacteria) in a non-dairy medium would provide some benefit to patients with lactose intolerance. According to the description in WO2004 / 089290 (Pharmachem), which addresses the issue of lactose intolerance, the use of dehydrated bacteria is not desirable. For example, on page 2 of WO2004 / 089290, it is considered that "Freeze-dried probiotic organisms, ie freeze-dried, are also available. Freeze-dried bacteria are in an abiotic state.

The need to moisten the microorganism before administration, in order to restore vitality, is a disadvantage since many bacteria do not survive rehydration. In addition, the surviving organisms, if any, are not immediately metabolically active, and cannot survive the extreme acidic conditions of the stomach. "Thus, according to WO2004 / 089290, the use of dry bacteria It does not offer a way forward.

Hove and others, Amer. J. Clin. Nutr, 1994, 59: 74-79, describe experiments carried out on subjects suffering from lactose malabsorption in which subjects were given capsules containing mixed lyophilized lactic acid bacteria containing B. bifidum , L. acidophilus, S, thermophilus and L. bulgaricus. Hove et al. Concluded that capsules containing lyophilized lactic acid bacteria do not relieve symptoms of lactose malabsorption (ie abdominal cramps, diarrhea and "meteorisms") and do not improve lactose hydrolysis in the intestine. thin in subjects with lactose malabsorption.

Therefore, in view of the descriptions described above, there is a need for an effective product that allows lactose intolerant subjects to consume food products that contain lactose - other than yogurts that contain live bacteria that are better tolerated by some of the sick - without suffering the symptoms usually associated with lactose intolerance.

Thus, an agent with a more reliable and / or better effect than the preparations containing only the enzyme lactase would thus satisfy an urgent need that exists for the many affected persons and would mean a considerable improvement and a great step forward in the Nutritional options available in the case of lactose intolerance.

Summary of the Invention

An objective of the present invention is to provide a more effective agent for use in the treatment or prevention of the symptoms of lactose intolerance and lactase deficiency, and to allow a subject to consume foods containing lactose even if the subject He suffers from lactose intolerance. In addition, it is an object of the invention to make it possible for those affected by lactose intolerance to eat food products that they were not allowed to eat until now, due to their lactose content. In addition, the objective is to provide an agent that can prevent, reduce or eliminate the appearance of symptoms of lactose intolerance after the intake of foods containing lactose or substances containing lactose.

The present inventors have found that the administration of the Lactobacillus delbrueckii ssp bulgaricus and Streptococcus thermophilus bacteria in dry form to subjects suffering from lactose malabsorption does not produce a statistically significant improvement in the digestion of lactose. This finding is consistent with the teachings in W02004 / 089290 and in Hove and others discussed above.

However, the present inventors have also found that compositions containing combinations of lactase and Lactobacillus delbrueckii ssp bulgaricus and / or Streptococcus thermophilus in a dry, uncoated form, when administered to subjects suffering from lactose malabsorption, are considerably more effective and reliable to improve lactose digestion, and exert their beneficial properties on a wider variety of subjects, than compositions containing only lactase.

Thus, contrary to what is suggested in the prior art discussed above, and particularly in US6008027 (Langner) and WO2004 / 089290 (Pharmachem), a non-dairy composition containing a combination of lactase and Lactobacillus delbrueckii ssp bulgaricus bacteria and / or Streptococcus thermophilus in a dry, uncoated form can provide an effective means to improve lactose digestion in subjects that poorly absorb lactose.

Thus, the present invention provides an effective agent to enable people suffering from lactose intolerance to consume lactose-containing food products without suffering the symptoms of lactose intolerance.

Accordingly, in a first embodiment (Modality 1.1), the invention provides a composition for use in the therapy of lactose intolerance or conditions resulting from lactase deficiency, wherein the composition is a solid dosage form not dairy comprising:

(to)

 a lactase which is stable and active in a pH range of 3 to 6; the term stable means that lactase

(to)

 it still has at least 30% of its activity after one hour at 37 ° C at a pH in the range of 3 to 6; Y

(b)

 one or more microorganisms that produce lactase and / or that contain lactase selected from Lactobacillus delbrueckii ssp bulgaricus and Streptococcus thermophilus; wherein the lactase (a) is different from a lactase derived from said one or more microorganisms (b); and wherein the one or more microorganisms that produce lactase and / or that contain lactase (b) are in dry form.

In additional embodiments the invention provides:

1.2 A composition for use in accordance with Modality 1.1 wherein the dosage form is different from a capsule coated with an enteric coating.

1.3 A composition for use according to Modality 1.1 wherein the organisms that produce lactase or that contain lactase are not enteric coated.

1.4 A composition for use according to any of Modalities 1.1 to 1.3 wherein lactase (a) is one that still has at least 30% of its activity after one hour at 37 ° C at a pH in the range of 4 to 6

1.5 A composition for use according to any of Modalities 1.1 to 1.3 wherein lactase (a) exhibits peak activity at a pH in the range of 3 to 6.

1.6 A composition for use according to any of Modalities 1.1 to 1.3 wherein lactase (a) exhibits peak activity at a pH in the range of 4 to 6.

1.7 A composition for use in accordance with any of Modalities 1.1 to 1.8 wherein the lactase (a) is derived from a microorganism, and the composition is substantially free of the microorganism from which the lactase is derived.

1.8 A composition for use according to any of Modalities 1.1 to 1.7 which contains Lactobacillus delbrueckii ssp bulgaricus and Steptococcus thermophilus.

1.9 A composition for use in accordance with any of Modalities 1.1 to 1.8 wherein the solid dosage form is selected from capsules, tablets, dragees, powders, tablets and granules.

1.10 A composition for use according to any of Modalities 1.1 to 1.9 which is in unit dosage form.

1.11 A composition for use according to Modality 1.10 wherein the unit dosage form is selected from capsules, tablets, dragees and sachets containing a powder, tablets or granules.

1.12 A composition for use according to Modality 1.11 wherein the unit dosage form is selected from capsules, tablets and dragees.

1.13 A composition for use in accordance with Modality 1.12 wherein the unit dosage form is a capsule.

1.14 A composition for use in accordance with any of Modalities 1.1 to 1.13 containing between 2000 and 12000 FCC units of lactase (a).

1.15 A composition for use in accordance with Modality 1.14 containing between 2000 and 6000 FCC units of lactase (a).

1.16 A composition for use in accordance with any of Modalities 1.1 to 1.15 containing between 50 million to 10 billion colony forming units of Lactobacillus bulgaricus and 50 million to 10 billion colony forming units of Steptococcus thermophilus.

1.17 A composition for use according to Modality 1.16 containing 100 million to 2 billion colony forming units of Lactobacillus bulgaricus and 100 million to 2 billion colony forming units of Steptococcus thermophilus.

1.18 A composition for use in accordance with any of Modalities 1.1 to 1.17 which is substantially lactose free.

1.19 A composition for use in accordance with any of Modalities 1.1 to 1.18 wherein Lactobacillus bulgaricus is Lactobacillus delbrueckii LB-VC18 (deposit number: DSM 23320).

1.20 A composition for use according to any of Modalities 1.1 to 1.19 wherein the Steptococcus thermophilus is Streptococcus thermophilus ST-VC18 (deposit number: DSM 23319).

1.21 A composition for use according to any of Modalities 1.1 to 1.20 wherein lactase (a) is derived from an Aspergillus species.

1.22 A composition for use according to Modality 1.21 wherein lactase (a) is derived from Aspergillus oryzae or Aspergillus niger.

1.23 A composition according to modality 1.22 wherein lactase (a) is derived from Aspergillus oryzae.

1.24 A composition for use in accordance with any of Modalities 1.1 to 1.23 where the

Composition is different from a composition that contains a combination of two or more herb extracts added together with exogenous lipase and exogenous protease.

1.25 A composition for use according to any of Modalities 1.1 to 1.24 wherein the composition contains (c) a dietary and pharmaceutically acceptable excipient or carrier.

1.26 A solid dosage form selected from capsules, tablets, dragees, powders, tablets and granules that is not coated by an enteric coating and that contains (a) a lactase that is stable and active in a pH range of 3 to 6; the stable term means that lactase (a) still has at least 30% of its activity after one hour at 37 ° C at a pH in the range of 3 to 6; and (b) one or more microorganisms that produce lactase or that contain lactase selected from Lactobacillus delbrueckii ssp bulgaricus and Steptococcus thermophilus; where (a) and (b) are as defined in any of Modalities 1.1 to 1.26; as long as the composition is different from a composition that collectively contains two or more herb extracts added together with exogenous lipase and exogenous protease.

1.27 A solid dosage form 1.26 which is selected from capsules, tablets and dragees.

1.28 A solid dosage form 1.26 which is selected from powders, tablets and granules and wherein said powders, tablets and granules are contained in an envelope.

1.29 A solid dosage form 1.27 which is a capsule.

1.30 A solid dosage form 1.27 which is a tablet.

1.31 A solid dosage form 1.27 which is a dragee.

Modalities and additional aspects of the invention

Additional embodiments and aspects of the invention will be apparent from the following description and examples.

The subject matter of the invention is an agent that can solve the problems described above in the introductory section of this application. The agent contains the enzyme lactase in combination with microorganisms. The microorganisms in question are lactic acid bacteria and particularly preferably Lactobacillus delbrueckii subspecies bulgaricus and / or Streptococcus thermophilus. A lactase within the meaning of this invention is an enzyme that is capable of converting (cleaving) lactose into glucose and galactose. Lactic acid bacteria within the meaning of this invention are particularly microorganisms that, through their own microbial lactase, can metabolize lactose and / or from which lactase can be released into the digestive tract of humans or animals. The microorganisms used in accordance with the present invention can preferably be lyophilized or spray dried (preferably lyophilized), in order to provide a longer shelf life and a good processing capacity.

Whenever Lactobacillus bulgaricus is used below, it refers to Lactobacillus delbrueckii subspecies (subsp.) Bulgaricus.

The agent according to the present invention can cause lactose cleavage in the food or in the food pulp in glucose and galactose after consumption. This means that lactose is no longer available for bacterial metabolism characterized by fermentation in the intestines.

The subject matter of the invention is therefore an agent that, with the aid of lactase in combination with microorganisms as defined in the claims appended hereto, reduces the bioavailability of lactose in the human or animal body.

The subject matter of the invention is also an agent which, with the aid of lactase in combination with microorganisms as defined in the claims attached herein, reduces the amount of lactose available to the human or animal body or to intestinal bacteria. colonized in it.

The subject matter of the invention is also an agent for use in the case of lactose intolerance containing lactase in combination with microorganisms as defined in the appended claims herein.

Lactase is an enzyme that has the property of converting lactose into glucose and galactose. While lactose is not absorbed in the small intestine, glucose and galactose are monosaccharides that are easily digestible and quickly absorbed. The lactases currently used in lactase preparations intended for oral use are called acid lactose (sometimes referred to as sour lactases) that are mostly produced using Aspergillus oryzae. Acid lactase is very effective in the acidic environment of the stomach after food consumption. Acid lactase is stable and active at a pH of around 3 - 6. Within the meaning of this application, the term "stable"

means that acid lactase still has at least 30% of its activity after one hour at 37�C and pH values of 3-6. The reference to 30% of its activity in the present context refers to the residual activity of the enzyme if, after the specified period of one hour, it is tested according to the acid lactase assay as described in the sixth edition of the Food Chemical Codex (FCC).

At a pH value of more than 6.5, as found in the small intestine, acid lactase is stable but no longer very active. Therefore, acid lactase can no longer develop a satisfactory effect on the small intestine. At present, lactase preparations therefore have to contain as much lactase as necessary to cleave the amounts of lactose that are usually consumed with food as much as possible in the stomach. A lactase that is stable and active at low pH values and that is effective in the acidic environment of the stomach and in the small intestine is not known or commercially available. Since the gastric transit time varies greatly depending on the food consumed and the amount of food consumed, the correct dosage of lactase preparations is often a problem for the user and therefore, as described above, users are frequently They are dissatisfied with the effect of these products. An increase in lactase activity per dose unit may be useful, but it has been found that the increase in effect and the reliability of the effect do not increase proportionally to the activity of lactase per dose unit. The timing of intake is also critical, since acid lactase is rapidly inactivated at the very low pH value (between 1 and 2.5) that prevails on an empty stomach. Thus, if a user consumes a lactase preparation too long (for example 10 minutes) before a meal on an empty stomach, it is possible that the enzyme has already irreversibly lost its activity at the time of food intake and then already It is not available for the desired cleavage of lactose, so that in this case the activity of lactase per dose unit does not play a role.

Certain lactic acid bacteria contain a lactase that is active and stable at the pH values that prevail in the small intestine. Lactic acid bacteria lactase, protected by the bacterial envelope, can at least partially (or to some extent) pass through the stomach into the small intestine, where two advantageous effects may come into play. On the one hand, microbial lactase can be released when the bacterial envelope is destroyed by digestive juices in the small intestine, so that lactose that has not yet been cleaved in the stomach by acid lactase is then cleaved by this lactase released by lactic acid bacteria. On the other hand, bacteria that remain intact can metabolize lactose and thus also contribute to a further reduction in the amount of lactose in the small intestine. Some people affected by lactose intolerance have a better tolerance to fermented milk products rather than unfermented milk products, which is partly due to the lactase content of lactic acid bacteria. However, the majority of those affected cannot consume fermented dairy products without problems. In studies in which lactic acid bacteria were administered to people with lactose intolerance, it was shown that they do not have good and reliable efficacy in relation to lactose digestion. This result has been demonstrated for a large number of different strains, in addition when used in a wide variety of different combinations. Therefore, in the case of lactose intolerance, the use of a preparation containing exclusively lactic acid bacteria does not make sense (see for example Hove and others as mentioned above). Considering that today the attempt is often made to add bacteria from the lactic acid to dairy products and other foods, which arrive intact to the large intestine in order to exert there beneficial effects on health, in the invention here It is preferred that the greatest possible number of lactic acid bacteria consumed be destroyed in the small intestine, so that its microbial lactase is released there, so that it is then available for lactose cleavage . By adding such bacteria from lactic acid, the efficiency and / or reliability of lactase preparations can be increased. Such preparation is able, on the one hand, to cleave lactose in the stomach (through acid lactase) and, on the other, to cleave lactose in the small intestine that is not cleaved in the stomach (by means of lactic acid bacteria lactase). Thus, by the addition of lactic acid bacteria as defined in the claims, a prolonged effect of the preparation is achieved, which leads to greater efficacy and / or reliability of the effect, which cannot be achieved purely by the Increase in lactase activity per unit dose or at least cannot be achieved profitably. In addition to the enzyme lactase, it is thus advantageous to add such microorganisms to the agent according to the present invention, which release their own lactase sufficiently active and stable in the small intestine and / or metabolize lactose in the small intestine. It has been found that Lactobacillus bulgaricus and Streptococcus thermophilus lactic acid bacteria are particularly suitable for this application, since these microorganisms predominantly, but at least partially, survive the stomach with an intact bacterial envelope that protects microbial lactase, but predominantly , but at least partially, they do not survive intact in the small intestine and thus release their microbial lactase there to a relevant extent.

Such a combined agent can be applied in the form of two separate dose units, for example in two separate tablets or capsules, of which one contains the lactase enzyme and the other the lactic acid bacteria.

Food products within the meaning of this invention are food products within the meaning of Regulation (EC) no. 178/2002 of the European Parliament and of the Council of January 28, 2002. Food products within the meaning of this invention particularly include food products for particular nutritional uses, foods for special medical purposes, food supplements, dietary supplements and food additives .

Food supplements in the sense of this invention are food supplements in the sense of

Directive 2002/46 / EC of the European Parliament and of the Council of June 10, 2002.

Food products for special nutritional uses within the meaning of this invention are food products for special nutritional uses within the meaning of Directive 2009/39 / EC of the European Parliament and of the Council of May 6, 2009 on food products intended for particular nutritional uses.

Foods for special medical purposes within the meaning of this invention are foods for special medical purposes within the meaning of Commission Directive 1999/21 / EC of March 25, 1999 on dietary foods for medical purposes special doctors.

This invention provides the enzyme lactase in combination with Lactobacillus bulgaricus and / or Streptococcus thermophilus for use in medicine, for example as a pharmaceutical composition. The subject matter of the invention is furthermore a product consisting of lactase in combination with Lactobacillus bulgaricus and / or Streptococcus thermophilus or containing these substances together with one or more other active ingredients, for use in a medical procedure. , particularly a procedure for the therapeutic treatment of the human or animal body.

The subject matter of the invention is also a composition for use in medicine, which contains a lactase and one

or more microorganisms that produce lactase and / or that contain lactase, selected from Lactobacillus bulgaricus and Streptococcus thermophilus, so that lactase preferably does not originate from the aforementioned microorganism (s). Within the meaning of this invention, a pharmaceutical composition is a product within the meaning of Directive 2001/83 EC in the version of Directive 2004/27 EC.

In accordance with a further aspect of the present invention, a food product containing lactase is provided in combination with lactic acid bacteria, particularly in combination with Lactobacillus bulgaricus and / or Streptococcus thermophilus. Furthermore, according to the present invention, a food product containing lactase in combination with lactic acid bacteria, particularly in combination with Lactobacillus bulgaricus and / or Streptococcus thermophilus, is provided in an amount effective to convert lactose into glucose and galactose

A food product can also be produced using a method in which lactase in combination with Lactobacillus bulgaricus and / or Streptococcus thermophilus is added to the food product in such a way that the effect of lactase and lactic acid bacteria begins predominantly after consumption of the food product. Such a food product tastes practically as an untreated food product and, as a result of the reduced lactose content that occurs after consumption, is suitable for consumption in the case of lactose intolerance. Predominantly within the meaning of this application it means that at least 50% of the cleavage effect of the enzyme lactose and lactic acid bacteria occurs after consumption of the food product. Alternatively, (a) at least 90%, (b) at least 80%, (c) at least 70% or (d) at least 60% of the effect may occur after consumption of the food product.

In accordance with a further aspect of the present invention, a medical device containing lactase is provided in combination with the lactobacillus bulgaricus and / or Streptococcus thermophilus lactic acid bacteria. The subject matter of the invention is also, consequently, a medical device containing lactase in combination with the lactobacillus bulgaricus and / or Streptococcus thermophilus bacteria, or containing it next to one

or more of other active ingredients.

A medical device within the meaning of this patent application is a medical device within the meaning of Directive 93/42 / EEC in the version modified by Directive 2007/47 / EC.

The various aspects of the invention are further described below. Whenever an agent is referred to below, it always also means a food product, medical device or pharmaceutical composition.

Lactase, specifically acid lactase, more specifically acid lactase derived from Aspergillus spp, even more specifically acid lactase derived from Aspergillus oryzae, has not yet been used in combination with Lactobacillus bulgaricus and / or Streptococcus thermophilus in the medical field Pharmaceutical, particularly in the case of lactose intolerance in humans or animals. Therefore, the invention presented here is the first medical indication for the combination of these substances.

The agents according to the present invention can be taken orally before meals, with meals or after meals, so that they can exert their cleavage effect on lactose in the pulp of food. Preferably, the agents according to the present invention are taken just before meals, during meals or immediately after meals. The agents according to the present invention may contain the enzyme and the bacteria of the lactic acid without additional additives. However, it is preferable that the agents according to the present invention also contain additives that are pharmaceutically acceptable and / or acceptable for food products, such as diluents, binders, stabilizers, preservatives, flavorings, etc. Such additives are commonly used and are well known for the production of pharmaceutical compositions, medical devices, food products, food products for particular nutritional uses,

food for special medical purposes, food supplements, dietary supplements and food additives, and the specialist in this field knows what additives and in what quantities are suitable for particular forms of presentation. Particularly preferably, the agents according to the present invention contain as additives dichloric phosphate, modified starch, microcrystalline cellulose, maltodextrin and / or Fibersol.

The agents according to the present invention can also be added to a food product before consumption. They can even be added to the food product in the production stage, in order to develop its effect at least partially but preferably predominantly after consuming the food product. This can be achieved by microencapsulation, for example. In this way, the usable lactose content of the food product would be reduced in a particularly advantageous manner. Therefore, preparations containing lactase in combination with Lactobacillus bulgaricus and / or Streptococcus thermophilus which predominantly do not release these substances until they enter the digestive tract of a human or animal especially in the stomach or small intestine are preferred. Therefore, the invention can be used for example in the production of milk and milk products, such as curd, yogurt, cream, cheese, pudding, milk drinks, mixed milk and ice cream drinks, and in the production of, for example chocolate and chocolate products, bakery products (for example cookies and cakes), breads, sweets, lactose-containing drinks, lactose-containing sauces (for example cream sauces) and lactose-containing sweeteners. For dishes that are boiled, baked or fried, the agents according to the present invention can, for example, be mixed in or sprayed on them after cooling. As far as milk and milk products are concerned, according to the present invention they contain acid lactase and at least one of the microorganisms selected from Lactobacillus bulgaricus and Streptococcus thermophilus.

The agents according to the present invention can also be added to a food product in order to exert its effect after consumption on the lactose from another food product. An example of this would be the addition of the agents according to the present invention to cereals, so that the reduction of lactose contained in milk occurs after the consumption of cereals prepared with milk, without damaging the taste of the milk

A subject matter of the invention are also agents that in addition to other active ingredients also contain lactase in combination with Lactobacillus bulgaricus and / or Streptococcus thermophilus.

The invention can be formulated in any form that is suitable for the intended route of administration. For oral administration, the agents according to the present invention are preferably formulated in the form of capsules (coated or uncoated), tablets (coated or uncoated), dragees (coated or uncoated), capsules containing tablets coated or uncoated, granules or micro or minitabletas, pressed tablets from coated or uncoated powder, tablets or micro or minitabletas. In addition, coated or uncoated gel capsules (soft gelatin capsules) are possible for oral administration.

The invention may also be presented as a dry or wet oral supplement. The formulation of the agents according to the present invention in powder form is particularly suitable for a mixture with a food product. The powder can be sprayed on a meal or it can be mixed in a pulp or a drink. It is particularly suitable if the agent offered as a bulk powder is packaged in single dosage amounts, such as in single bags, or if it is provided in a dosing apparatus.

It is especially preferable if the agent according to the present invention is formulated as a powder or as granules or tablets in capsules or as a tablet that is administered orally.

The agents or compositions of the invention typically contain at least one dietary and pharmaceutically acceptable carrier or excipient.

For oral administration, the active ingredients may be contained in acceptable carriers and / or excipients. The term "acceptable carrier" refers to a carrier that delivers the active ingredient to its destination site and does not cause significant damage to the recipient, human or animal. However, the exact form of the carrier is not important.

A carrier can be, for example, a capsule wrap, an envelope or a matrix into which the active ingredients are enclosed.

The total amount of the carrier and / or excipient in relation to an agent according to the present invention is preferably between 5 and 99.9% by weight, more preferably between 10 and 95% by weight and even more preferably between 25 and 90% by weight of the composition.

Suitable excipients and / or carriers include maltodextrin, calcium carbonate, dichloric phosphate, tricyclic phosphate, microcrystalline cellulose, dextrose, rice flour, magnesium stearate, stearic acid, sadic croscarmellose, sadic starch glycolate, crospovidone, sucrose, vegetable resins, lactose, methyl cellulose, povidone, carboxymethyl cellulose, corn starch, modified starch, fibersol, gelatin, hydroxypropyl methylcellulose and the like (which

include mixtures of these). Preferable carriers include calcium carbonate, magnesium stearate, maltodextrin, dichloric phosphate, modified starch, microcrystalline cellulose, fibersol, gelatin, hydroxypropyl methylcellulose and mixtures thereof. The various ingredients and the excipient and / or carrier are mixed and formed in the desired form using common methods. The presentation form that is intended for oral administration according to the present invention, such as a tablet or capsule, can be coated with a coating that is resistant to low pH values or a gastro-resistant coating. In addition, a coating that is not resistant to low pH values but that provides delayed release of the respective enzyme or bacterium from the respective lactic acid at low pH values can be used. It is also possible to prepare the agent according to the present invention as tablets, granules, or coated micro or minitabletas (see above) that can be filled in uncoated capsules or that can be pressed into uncoated tablets. Suitable coatings are, for example, cellulose acetate phthalate, cellulose derivatives, shellac, polyvinylpyrrolidone derivatives, acrylic acid, polyacrylic acid derivatives and polymethyl methacrylate (PMMA), such as Eudragit (de R �hm GmbH, Darmstadt), particularly Eudragit FS30D and Eudragit L30D-55. By adding, for example, sodium hydroxide solution to the coating agent, the pH resistance of this coating agent can also be influenced. Additional details about formulation and administration methods can be found in the 21st edition of "Remington: The Science & Practice of Pharmacy", published in 2005 by Lippincott, Williams & Wilkins, Baltimore, United States, and in the "Lehrbuch der Pharmazeutischen Technologie "by Professor Bauer, 18th edition, published in 2006 by Wissenschaftliche Verlagsgesellschaft (ISBN 3804-72222-9), so that the documents are incorporated in this way as a reference.

Although in accordance with the present invention such lactic acid bacteria are preferably used to survive the stomach with its bacterial envelope intact as much as possible, however it may be advantageous to apply a gastro-resistant coating to the lactic acid bacteria or formulate them. , for example, in capsules, dragees, tablets, tablets, granules or crystals with a gastro-resistant coating, so that a proportion of bacteria as large as possible reaches the small intestine or at least the duodenum (the upper part of the small intestine). The bacteria of the lactic acid may also be contained in, for example, tablets, granules with a gastro-resistant coating, or a gastro-resistant coating may be applied to the powder of the bacteria of the lactic acid (for example using a spray method). Tablets, granules or powder can be filled in uncoated capsules. These may also contain the lactase that is supposed to be released in the stomach. A coating to protect the bacteria from lactic acid against gastric acid would be very advantageous particularly if you use lactic acid bacteria whose bacterial envelope would otherwise be destroyed in the stomach, so that your microbial lactase will be inactivated. a in the stomach due to for example the low pH value of there.

Other carriers and / or dietary and pharmaceutically acceptable excipients suitable for use in the present invention include water, mineral oil, ethylene glycol, propylene glycol, lanolin, glyceryl stearate, sorbitan stearate, isopropyl myristate, isopropyl palmitate, acetone, glycerin, phosphatidylcholine, sodium cholate or ethanol, but not limited to these.

Compositions for use in the present invention may further comprise at least one coemulsifying agent, which includes oxyethylenated sorbitan monostearate, fatty alcohols, such as stearyl alcohol or cetyl alcohol, or esters of fatty acids and polyols, such as glyceryl stearate, but not limited to these.

Preferably, the agents used in the present invention are provided in stabilized form. Generally, the stabilization methods and methods that can be used in accordance with the present invention include any and all methods for the stabilization of chemical or biological material known in the art, comprising for example the addition of agents chemical methods that are based on temperature modulation; the methods that are based on irradiation or combinations thereof. Chemical agents that can be used in accordance with the present invention include, among others, preservatives, acids, bases, salts, antioxidants, viscosity improvers, emulsifying agents, gelatinizers, and mixtures thereof.

Acid lactase is commercially available (for example, Amano Japan and DSM Netherlands) and is conventionally prepared in a microbiological manner with the help of the Aspergillus oryzae microorganism. Lactobacillus bulgaricus and Streptococcus thermophilus are also commercially available (eg Danisco, Denmark, or Christian Hansen, Denmark). However, the invention is not limited to the acid lactose enzymes that are commercially available at the moment, but generally refers to enzymes that can cause the conversion of lactose - specifically or unspecifically - to glucose and galactose in combination with lactic acid bacteria that can break down lactose in the small intestine, that is, either by the release of lactase or through the intracellular metabolization of lactose.

Lactase activity is defined in accordance with the present invention in FCC units (Food Chemical Codex units). An enzyme unit releases an o-nitrophenol micromol per minute at 37 ° C and a pH of 4.5, under the conditions described in the sixth edition of the Food Chemical Codex (see the complete specification of the measurement, the description of the test, the calculation of the activity).

At an enzyme activity determined according to this definition, the agent according to the present

The invention should contain 500 to 50,000 FCC units, preferably 2,000 to 20,000 FCC units and particularly preferably 3,000 to 12,000 FCC units per unit dose of lactase.

If the amounts below are considered (in colony forming units, abbreviated as cfu) with respect to the lactic acid bacteria contained in the agent according to the present invention, they refer to the amount of l acidic acid bacteria. It is still viable (in cfu) at the end of shelf life of the agent according to the present invention. It should be noted that preparations containing lactic acid bacteria are preferably stored in a dry and cool place. Thus, in order to achieve a sufficiently prolonged viability of the lactic acid bacteria, it may be advantageous to store the agent according to the present invention in an airtight container and preferably impermeable to water vapor (for example of glass or aluminum), equipped with a desiccant (for example in the lid), for example silica gel or molecular sieves.

The amount of lactic acid bacteria is considered in cfu (colony forming units). The agent according to the present invention should contain between 10 million and 200 billion (billion = billion) of cfu, preferably between 50 million and 10 billion cfu and particularly preferably between 100 million and 2 trillion cfu (for example between 100 million and 1 trillion cfu) per strain of lactic acid bacteria.

In the case of a combined agent containing Lactobacillus bulgaricus and / or Streptococcus thermophilus in addition to the enzyme lactase, a dose unit should contain between 10 million and 200 billion cfu of Lactobacillus bulgaricus, preferably between 50 million and 10 trillion cfu of Lactobacillus bulgaricus and particularly preferably between 100 million and 2 billion cfu (for example between 500 million and 2 billion cfu) of Lactobacillus bulgaricus. In addition to or instead of the Lactobacillus bulgaricus bacteria, the agent according to the present invention should contain between 10 million and 200 billion cfu of Streptococcus thermophilus, preferably between 50 million and 10 trillion cfu of Streptococcus thermophilus and particularly preferably between 100 million and 2 billion cfu (for example between 500 million and 2 trillion cfu) of Streptococcus thermophilus per unit dose.

An agent according to the present invention has proved particularly advantageous, which contains between 2000 and 12000 FCC units (for example between 2000 and 6000 FCC units) of acid lactase, between 100 million and 10 billion cfu (for example between 100 million and 2 billion cfu) of Lactobacillus bulgaricus and between 100 million and 10 trillion cfu (for example between 100 million and 2 billion cfu) of Streptococcus thermophilus per dose unit. Particularly advantageous results can be achieved if the acid lactase is combined with the two strains of Lactobacillus delbrueckii LB-VC18 (deposit number: DSM 23320) and Streptococcus thermophilus ST-VC18 (deposit number: DSM 23319) deposited in the Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (German Collection of Microorganisms and Cell Cultures) in Braunschweig, Germany.

The wide variety of the above mentioned dosages can be explained by the fact that the agent according to the present invention can be applied in a wide variety of different severity levels and also in milder lactose metabolism disorders. Affected humans and / or animals react to a certain degree of lactose to a different extent. Apart from this, the different dosages result in addition to the fact that very variable amounts of lactose are supplied to the body, depending on the respective food consumed.

The agent according to the present invention may contain, for example, between 500 and 50,000 FCC units (for example between 500 and 30,000 FCC units) of lactase and between 10 million cfu and 200 billion cfu of Lactobacillus bulgaricus and / or between 10 million cfu and 200 billion cfu of Streptococcus thermophilus per unit dose. In addition to one or both of these microorganisms, the agent according to the present invention may in each case also contain between 10 million cfu and 200 billion cfu of the following lactic acid bacteria: Lactobacillus delbrueckii (subspecies other than Lactobacillus bulgaricus), Lactobacillus acidophilus, Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus paracasei, Lactobacillus brevis, Lactobacillus plantarum, Lactobacillus salivarius, Lactobacillus rheuteri, Lactococcus lactis, Pediococcos acidilacti, Bifidobacterium lactis, Bifidobacterium bifidum, Bifidobacterium breve, Bifidobacterium longum, Bifidobacterium infantis.

Apart from this, suitable additives can be used in the required quantities.

The invention can be made available for medical and non-medical purposes, for example as a pharmaceutical composition, a medical device or a food product.

Long-term therapy in the present context has to be understood in a broad sense. Within the meaning of this application, therapy not only refers to giving therapy to or treating diseases, but also very generally includes the use of the agent according to the present invention or of the agents according to the present invention in the intolerance to lactose and lactase deficiency in humans or animals (since in almost all mammals the ability to digest lactose decreases after stopping breastfeeding). Within the meaning of this application, the therapy accordingly encompasses in particular the use of the agent according to the present invention or of the agents according to the present invention to prevent, eliminate, relieve, reduce or act as a preventive measure against the onset of (a) all types and levels of severity of symptoms, (b) all types and levels of severity of

indispositions and (c) all forms of health damage and discomfort that may occur as a result of ingestion of lactose or food products containing lactose or through the release of lactose in the digestive tract of humans or animals from other substances, for example pharmaceutical compositions. In addition, within the meaning of this request, the therapy encompasses eradication, relief, elimination, reduction, preventive measures against and prevention of abdominal distension, gas, stomach pain, stomach pain with colic, colic , diarrhea, watery diarrhea, intestinal noises, abdominal cramps, belching, nausea, vomiting, flatulence, microbiological imbalance in the intestines or increased intestinal gas production associated with ingestion of lactose or food products containing lactose or with Lactose release in the digestive tract of humans or animals from other substances, for example pharmaceutical compositions. In addition, within the meaning of this application, the therapy includes eradication, relief, elimination, reduction, preventive measures against and prevention of any type of depression, headaches, migraines, eczema, rashes cutaneous and, lack of appetite and weight loss associated with the ingestion of lactose or food products containing lactose or with the release of lactose in the digestive tract of humans or animals from other substances, for example pharmaceutical compositions or associated with insufficient digestion of lactose or malabsorption of lactose. In addition, therapy within the meaning of this invention encompasses the reduction of the bioavailability of lactose in the human or animal body and the reduction of the amount of lactose available to the human or animal body or to the intestinal bacteria that colonize therein. .

In the context of this application the term therapy includes prophylaxis and treatment.

Examples

The invention will now be illustrated, but not limited, with reference to the examples set forth below.

Example 1

Capsule compositions

The following capsule compositions can be prepared by mixing Aspergillus oryzae lactase, and lyophilized Lactobacillus bulgaricus and Streptococcus thermophilus.

Composition in capsule A - Capsule size 1

One size 1 gelatin capsule (0.5 ml volume) that can be obtained from Capsugel BVBA, Bornem, Belgium is filled with 70 mg lactase (lactase activity is 50,000 FCC / g), 40 mg Streptococcus thermophilus (30 billion of cfu / g) lyophilisate, 30 mg of Lactobacillus bulgaricus (50 billion cfu / g) lyophilisate and 200 mg of dicyclic phosphate per capsule.

A preferred strain of Streptococcus thermophilus is Streptococcus thermophilus ST-VC18 (deposit number: DSM 23319) and a preferred strain of Lactobacillus bulgaricus is Lactobacillus delbrueckii LB-VC18 (deposit number: DSM 23320).

Composition in capsule B - Capsule size 3

One size 3 jelly capsule (volume 0.3 ml) that can be obtained from Capsugel BVBA, Bornem, Belgium is filled with 70 mg lactase (lactase activity 50,000 FCC / g), 30 mg Streptococcus thermophilus (30 billion cfu / g), 20 mg of Lactobacillus bulgaricus (50 billion cfu / g) and 100 mg of dichloric phosphate per capsule.

Composition in capsule C - Capsule size 0

One size 2 jelly capsule (0.68 ml volume) that can be obtained from Capsugel BVBA, Bornem, Belgium is filled with 150 mg lactase (lactase activity 50,000 FCC / g), 100 mg Streptococcus thermophilus (30 billion cfu / g), 50 mg of Lactobacillus bulgaricus (50 billion cfu / g) and 250 mg of dicyclic phosphate.

Composition in capsule D - Capsule size 2

A 3-size gelatin capsule (0.37 ml volume) that can be obtained from Capsugel BVBA, Bornem, Belgium is filled with 100 mg lactase (lactase activity 100,000 FCC / g), 50 mg Streptococcus thermophilus (30 billion of cfu / g), 30 mg of Lactobacillus bulgaricus (50 billion cfu / g) and 100 mg of dichloric phosphate.

Example 2

An investigation of the effect of a combination of lactase and lactic acid bacteria on the digestion of lactose in subjects that poorly absorb lactose

An investigation was carried out to compare the effects of lactase, lyophilized lactic acid bacteria and

a combination of lactase and lactic acid bacteria on the digestion of lactose in subjects that poorly absorb lactose.

The research consisted of a randomized, double-blind, placebo-controlled cross-sectional study that involved twenty-four human subjects who poorly absorb lactose. Thirty subjects were initially recruited for the study but it was found that six did not absorb lactose poorly and therefore the results obtained from these six subjects were excluded from the analysis of the results.

The study includes an entrance exam and five days of experimentation, separated by 4 washing periods of two weeks each. During the study, participants were asked not to change their eating habits and refrain from taking vitamins, minerals and other supplements. At the beginning of each experimental day, after an overnight fast, a first breath sample was taken from each subject. After that each subject ingested one of the five test preparations in random order.

Test preparations were provided in capsules of identical appearance, of which one was administered per experimental meal to the study participants along with 150 ml of milk to which 5 g of lactose was added, resulting in a lactose fortified milk containing approximately 12.5 g of lactose. The compositions of the five test preparations are as follows:

Preparation (a): Capsule containing 1 x 109 cfu of Streptococcus thermophilus plus 1 x 109 cfu of Lactobacillus delbruckii ssp. bulgaricus

Preparation (b): containing 3300 FCC of Aspergillus oryzae acid lactase

Preparation (c): Capsule containing 9000 FCC of Aspergillus oryzae acid lactase

Preparation (d): Capsule containing a combination of 3300 FCC of acid lactase plus 1 x 109 cfu of Streptococcus thermophilus and 1 x 109 cfu of Lactobacillus delbruckii ssp. bulgaricus (test preparation a))

Preparation (e): Dichloric Phosphate (placebo).

The bacteria of lactic acid in test preparations (a) and (d) are the ST-VC 18 and LB-VC18 strains described in Example 1 and in the description on page 24 above. The test preparation (b) is Lactrase 3300 (Pro Natura Gesellschaft f�r Ern�hrung gesunde mbH, Bad Vilbel, Germany), the test preparation (c) is Lactrase Plus 9000 (Pro Natura Gesellschaft f�r Ern�hrung gesunde mbH, Bad Vilbel, Germany) and the test preparation (d) is Lactrase 3300 (Pro Natura) to which the lactic acid bacteria were added.

The randomization of the participants in the study as well as the packaging and coding of the test preparations was carried out by the subjects who were not involved in the study in any other way.

The dose of lactose that was given (12.5 g) is lower than for a classic lactose malabsorption test (50 g), but it is closer to the lactose content of normally ingested food products. In the classic lactose malabsorption test (carried out with 50 g of lactose) the peak H2 concentrations of 20 ppm and more are considered a lactose malabsorption test. However, due to the smaller doses of lactose that were given in the present study, a lower threshold was used to determine if a participant is a subject that poorly absorbs lactose. Therefore, participants with peak H2 concentrations of 13 ppm and more with placebo were considered to absorb lactose poorly and the results obtained from these participants were included in the study.

The effect of test preparations on lactose digestion was measured using H2 breath tests that were carried out on a MicroLyzer Model 12i, Quintron Instruments, Milwaukee, WI 53215, United States. When the bacteria in the colon ferment carbohydrates, hydrogen gas is released in approximately 90% of the population. Therefore, the amount of hydrogen in the exhaled breath provides a good indicator of the amount of lactose that is fermented in the colon and is also a good indicator of lactase deficiency in the small intestine.

Breath samples were taken for 3 hours. The peak H2 concentration was chosen as a parameter for lactose digestion. The peak H2 concentrations for each of the subjects and each of the five preparations are shown in the table below.

H2 max values

Subject id

Preparation (e) Preparation (a) Preparation (b) Preparation (c) Preparation (d)

3501

 27 48 2 16 7

3503

 fifteen 12 4 2 18

3504

 37 36 33 46 9

3505

 27 10 24 16 fifteen

3506

 27 14 29 35 18

3507

 48 44 25 14 7

3508

 18 38 24 2. 3 fifteen

3509

 22 33 Four. Five 27 twenty

3510

 33 33 28 33 28

3511

 29 29 12 Four. Five 18

3513

 13 10 6 4 17

3514

 17 9 8 10 4

3516

 fifteen twenty-one 7 10 13

3517

3. 4 19 10  3 10

3518

fifteen  5 16 22 eleven

3520

fifteen  7 14 10 6

3521

 27 12 31 5 9

3522

 13 5 8 2 16

3524

 twenty-one 16 5 3 5

3526

 26 18 16 16 19

3527

 19 17 18 16 12

3528

 22 fifteen 18 12 13

3529

 Four. Five Four. Five 61 24 fifteen

3530

 53 33 22 39 43

The statistical analysis of the results of the study showed that there was a significant overall effect among the diets with respect to the maximum H2 values obtained (Friedmann's test, p = 0.001). The multiple comparison (Wilcoxon) showed significant differences with respect to the maximum H2 values between the preparation (b), which contains 3300 FCC units of lactase, and the placebo preparation (e), between (c), which contains 9000 units FCC of lactase, and the placebo preparation (e) and between the combination preparation (d), which contains 3300 FCC of lactase plus 2 billion cfu of lactic acid bacteria, and the placebo preparation (e). In addition, the average variation (dispersion) of the maximum H2 values, that is, the area of 25/75% interquartile for the preparation (d) was only half the size of the average variations of the other test preparations (ie say the preparations (a), (b) and (c)).

The results showed that the four preparations containing active ingredient (s); that is the preparation

(a) (bacteria alone), preparation (b) (3300 FCC), preparation (c) (9000 FCC) and preparation (d) (combination preparation) reduced H2 concentrations of breath at least in some subjects but only the 3300 FCC preparation (preparation (b)), the 9000 FCC preparation (preparation (c)) and the combination preparation (preparation (d)) showed a significant effect. The combination preparation (d) was even more effective than the preparation containing 9000 FCC (c) and also showed a significantly lower average variation in the exhalation values of H2. An analysis of the data showed that fewer participants who received the combination preparation (d) enjoyed little or no beneficial effect. The advantage of the combination preparation (d) therefore was not only its strongest contribution to the digestion of lactose but also that it was effective in a larger proportion of the study participants. Therefore it can be concluded that more subjects who poorly absorb lactose would benefit from the digestive effect of lactose from the combination preparation (d) than those who would benefit from the individual preparation of lactic acid bacteria ( a) and lactase preparations (b) and (c).

Example 3

An investigation of lactose turnover by incubating lactose with Streptococcus thermophilus and Lactobacillus bulgaricus bacteria

In this study, the degree of lactose cleavage by lyophilized bacteria and resuspended cultures of Streptococcus thermophilus and Lactobacillus bulgaricus was investigated.

The Streptococcus thermophilus strain used was Streptococcus thermophilus ST-VC18 (deposit number: DSM 23319) and the Lactobacillus bulgaricus strain used was Lactobacillus delbrueckii LB-VC18 (deposit number: DSM 23320).

Lyophilisates from both cultures were resuspended in phosphate buffer, pH 7.3, both separately and in a 1: 1 mixture of both bacteria based on colony forming units (cfu). In addition, investigations were made as to whether lactase activity in these bacterial preparations is stable in an acidic medium comparable to that of the stomach (simulated gastric fluid = SGF). In all cases, lactose cleavage was tested in the presence of pancreatic enzymes (containing Kreon capsules, 10,000 units of lipase, 8,000 units of amylase and 600 units of pancreas protease according to the European Pharmacopoeia and sodium dodecyl sulfate ) from porcine pancreas containing enzymes of the active pancreas plus detergent to mimic the activity of bile (simulated intestinal fluid = SIF). The amount of glucose, as measured enzymatically by the

glucose oxidase assay, was taken as a measure of lactose turnover. The glucose oxidase assay was used as described and as referred to in the Sigma Aldrich Chem. Co., 1996 glucose oxidase assay data sheet.

Freeze-dried bacteria (1 mg / ml) were resuspended in 1 ml of phosphate buffer (0.1 mol / l). Lactose (10 mmol / l) and Kreon were added at a concentration of 2 capsules per liter (recommended intake of 1 to 2 capsules per meal) and the suspension was incubated for 10 and 30 minutes. In this experiment, incubation of Lactobacillus bulgaricus resulted in the release of 1.7 mmol / I of glucose after 30 minutes and incubation with Streptococcus thermophilus produced 2.8 mmol / I of glucose, indicating changes of 17 and 28% respectively.

In order to test the acid stability of lactase activity, resuspended lyophilisates (1 mg / ml of bacterial lyophilisate per ml of suspension) were incubated with simulated gastric fluid, pH 4.0, for 10, 30 and 60 minutes. The suspension was neutralized by the addition of phosphate buffer to give a pH of approximately 7.0, and then lactose (10 mmol / l) and Kreon were added and lactase activity was determined as described above (30 min incubation). �n of the bacterial suspension with lactose, ie identical conditions).

Lactobacillus suspensions released 2.0 to 2.5 mmol / l of glucose after 30 minutes and Streptococcus suspensions released 3 to 3.5 mmol / l glucose after this period. No differences were observed between the samples incubated for 10, 30 or 60 minutes with simulated gastric fluid.

This experiment demonstrates that lactase activity in both preparations is stable in a reconstituted stomach environment.

In a third experiment, lyophilisates of Streptococcus thermophilus and Lactobacillus bulgaricus were mixed on a 1: 1 basis (based on the number of colony forming units). For replacement measurements, 1 mg of Streptococcus bacteria and 0.6 mg of Lactobacillus were resuspended in 1 ml of phosphate buffer and the resulting mixture was incubated with simulated intestinal fluid. After incubating this mixture for 30 minutes with 10 mmol / I lactose, 3.1 mmol of glucose was detected.

The same mixture was incubated for 30 minutes with simulated gastric fluid before lactase determination. After neutralization and incubation for 30 minutes in intestinal fluid simulated with 10 mmol / l lactose, the glucose concentration was found to be 3.15 mmol / I thereby demonstrating the stability of this mixture against the conditions here. dicas

Experiments demonstrate the presence of lactase activity in lyophilized bacteria. This lactase activity is stable and active in simulated intestinal fluid, even after undergoing simulated gastric fluid before being tested in the SIF. From these experiments it can be concluded that in vivo hydrolysis of lactose in the small intestine can be achieved by ingesting the combination of both bacteria or by ingesting each bacterium itself and that there is no significant difference between the activities of individual types of bacteria when used alone or when used in combination.

The data provided in the previous examples demonstrate that the combinations of acid lactase and Streptococcus thermophilus and / or Lactobacillus bulgaricus bacteria provide a more effective and more reliable improvement in lactose digestion in subjects that poorly absorb lactose than the compositions containing only lactase or compositions containing only bacteria. Furthermore, contrary to the teachings in the prior art documents discussed above, it is possible to present the bacteria in dry form and it is not necessary to coat the bacteria with an enteric coating in order to achieve the desired therapeutic effect.

Claims (18)

 Claims 1. A composition for use in the therapy of lactose intolerance or conditions resulting from lactase deficiency, wherein the composition is a non-dairy solid dosage form comprising:

(to)

a lactase which is stable and active in a pH range of 3 to 6; the stable term means that lactase (a) still has at least 30% of its activity after one hour at 37 ° C at a pH in the range of 3 to 6; Y

(b)

one or more microorganisms that produce lactase and / or that contain lactase selected from Lactobacillus delbrueckii ssp bulgaricus and Steptococcus thermophilus; wherein the lactase (a) is different from the lactase derived from said one or more microorganisms (b); and wherein the one or more microorganisms that produce lactase and / or that contain lactase (b) are in dry form.

2. A composition for use according to claim 1 wherein the dosage form is different from a capsule coated with an enteric coating.

3.

A composition according to claim 1 wherein the organisms that produce lactase or that contain lactase are not enteric coated.

Four.

A composition for use according to any one of claims 1 to 3 wherein the lactase (a) exhibits peak activity at a pH in the range of 3 to 6.

5.

A composition for use according to any one of claims 1 to 4 wherein the lactase (a)

25 is derived from a microorganism, and the composition is substantially free of the microorganism from which lactase is derived.

6.

A composition for use according to any one of claims 1 to 5 which contains Lactobacillus delbrueckii ssp bulgaricus and Steptococcus thermophilus.

7.

A composition for use according to any one of claims 1 to 6 wherein the solid dosage form is selected from capsules, tablets, dragees, powders, tablets and granules.

8. A composition for use according to any one of claims 1 to 7 which is in unit dosage form.

9.

A composition for use according to claim 8 wherein the unit dosage form is selected from capsules, tablets and dragees.

10.

A composition for use according to claim 9 wherein the unit dosage form is a capsule.

eleven.

A composition for use according to any one of claims 1 to 10 containing between 2000 and 12000 FCC lactase enzyme units (a).

12.

A composition for use according to any one of claims 1 to 11 containing 50 million to 10 billion colony forming units of Lactobacillus bulgaricus and 50 million to 10 billion colony forming units of Steptococcus thermophilus.

13.

A composition for use according to any one of claims 1 to 12 which is substantially lactose free.

14.

A composition for use according to any one of claims 1 to 13 wherein Lactobacillus bulgaricus is Lactobacillus delbrueckii LB-VC18 (deposit number: DSM 23320).

fifteen.

A composition for use according to any one of claims 1 to 14 wherein the Steptococcus thermophilus is Streptococcus thermophilus ST-VC18 (deposit number: DSM 23319).

16.

A composition for use according to any one of claims 1 to 15 wherein the lactase (a) is derived from a species of Aspergillus.

17.

A composition for use according to claim 16 wherein the Aspergillus species is Aspergillus oryzae.

A composition for use according to any one of claims 1 to 17 wherein the composition is different from a composition containing a combination of two or more herb extracts added together with exogenous lipase and exogenous protease .

19.

A composition according to any of claims 1 to 18 which contains (c) a dietary and pharmaceutically acceptable excipient or carrier.

twenty.

A solid dosage form selected from capsules, tablets, dragees, powders, tablets and granules that is not coated by an enteric coating and that contains (a) a lactase that is stable and active in a pH range of 3 to 6; the stable term means that lactase (a) still has at least 30% of its activity after one hour at 37 ° C at a pH in the range of 3 to 6; and (b) one or more microorganisms that

10 produce lactase or containing lactase selected from Lactobacillus delbrueckii ssp bulgaricus and Steptococcus thermophilus; wherein (a) and (b) are as defined in any one of claims 1 to 19; as long as the composition is different from a composition that collectively contains two or more herb extracts added together with exogenous lipase and exogenous protease. 21. A solid dosage form according to claim 20 which is selected from capsules, tablets and dragees.